PROJECT SUMMARY/ ABSTRACT Approximately 30% of the world?s population is now classified as obese or overweight; this high incidence is associated with increased risk of cardiovascular disease, cancer and diabetes, and reflects a worldwide healthcare crisis. At this time new therapies are needed urgently. The broad, long-term objectives of this project are to validate a new paradigm for drug discovery efforts in this area. A major emphasis in drug discovery today involves targeting changes in signal transduction pathways that are critical drivers of a disease state, so as to address specifically the etiology of that disease. Insulin and leptin trigger tyrosine phosphorylation-dependent signaling pathways that control appetite and feeding, energy expenditure and glucose homeostasis. In fact, diabetes and obesity are diseases of insulin- and leptin-resistance, respectively. This drew attention to those members of the family of protein tyrosine phosphatases (PTPs) that would normally attenuate these signaling pathways as potential therapeutic targets. Pharmacological inhibition of these PTPs would be expected to promote signaling and help to overcome the resistant state. Major programs in industry focused on developing small molecule PTP inhibitors, but they have been frustrated by technical challenges arising from the chemical properties of the PTP active site. As a result, industry classifies PTPs as potentially ?non-druggable? and they remain a largely untapped resource for drug development. Nevertheless, the extensive biological data validating phosphatases, such as PTP1B, as therapeutic targets, emphasizes the importance of discovering new approaches to inhibitor development that circumvent the problems encountered with active site-directed inhibitors. Therein lies the opportunity for academia, with its freedom to think without boundaries and to challenge existing paradigms. It is perhaps only in academia that new and innovative approaches, such as described in this proposal, can be taken to develop strategies that exploit areas and targets currently overlooked by industry, thereby benefiting patients through discovery of new therapies. This proposal, which builds on the foundation of progress during previous funding periods, addresses this problem from a unique and innovative perspective. The emphasis remains on defining and harnessing a novel mechanism for physiological regulation of PTP function by reversible oxidation in response to stimuli, such as insulin and leptin. In the previous funding period, we characterized scFv antibodies that recognize and sequester the oxidized, inactive form of PTP1B, promoting enhanced and sustained insulin signaling. In addition, we provided proof of concept for small molecule inhibitors that mimic the effects of these antibodies. In this competing renewal, addressing the following Specific Aims will develop further this unique approach to drug development for the treatment of diabetes and obesity: 1: To define non-catalytic functional and regulatory events mediated by oxidized PTPs. 2: To validate TCPTP-OX as a therapeutic target for diabetes and obesity. 3: To develop an alternative approach to trapping PTPs in their reversibly oxidized, inactive state. 4: To test PTP-OX stabilizers on insulin and leptin signaling in animal models of diet-induced and syndromic obesity.